In recent years, a striking progress has taken place in the studies and application of human fetal cadaver tissues. A completely new area of therapy is being developed, i.e. cell therapy that permits to fill an insufficient functional activity of damaged and sore tissues through the application of medicinal preparations based on fetal cell suspensions and prepared from fetal tissues. By using cell suspensions prepared from fetal embryo tissues, it is possible to attain such conditions where embryo cells administered to the organism of a patient may survive, locate their respective target organs and produce descendants, thereby filling deficient functional units of organs and tissues.
Embryo cells are often capable of migrating, establishing intercellular links, proliferating, differentiating, and responding to effects. They can produce considerable amounts of biologically active substances, e.g. hemopoietic growth factors, interleukins, nerve growth factor, allogenic and neurotrophic factors etc. Embryo cells cause a weaker immune response than mature cells due to the late expression of the main antigens in the process of their maturation. Embryo cell suspensions contain much lower amounts of highly active cells such as leukocytes, endothelium of vessels, dendritic cells etc. During the early fetal period transplants do not have mature lymphocytes, and are tolerant to recipient's tissues.
The vital potential of various Fetal Cells and Tissues generate curative effects. Main of these effects are: recovery the suppressed hematopoiesis (normalization of the quantity of erythrocytes, leukocytes, lymphocytes, thrombocytes); stimulation, correction or suppression of the immune system; normalization of homeostasis (glycemia, lipidemia, creatinemia, mineral and water metabolism, blood pressure etc.); stimulation of trophic functions.
We believed that the most expedient way of initiation our effort would comprise a model of AIDS in which various kinds of injuries to numerous systems of the human organism are manifested. In addition, penetrating of the U.S. medical market by medicinal preparation based on Fetal Cells would be the most prospective under the flag of treatment of AIDS which constitutes s considerable menace to the U.S. society and requires urgent actions aimed at saving a great number of people suffering from this disease.
We studied the impact produced by Petal Cells and revealed their powerful curative effect on various functions of organism in patients suffering from AIDS. The methods of AIDS treatment with the use of Fetal Cells, invented by the author and contributors, has been protected by U.S. Pat. No 5,811,069 [14].
However, the about mentioned application addressed not only a method of treatment but also a medicinal preparation that would combine all the advantages related to the use of living cells with convenient way of handling, intrinsic in common medicinal preparation. We managed to prepare such preparations comprising the subject of this invention. During several years we have been using it successfully in our specialized Clinic of Cell Therapy of the National Medical University, as well as in process of providing assistance to severely sick people in medical institutions of 15 reputable research institutes and specialized centers of the Academy of Medical
Science and the Ministry of public health of Ukraine in an area of treatment of numerous diseases characterized by injuries to the immune system, metabolism, and hemopoiesis. Substantiation of application of this medicinal preparation and relevant clinical examples are given hereinafter with reference to patients suffering from AIDS, wherein the inventive medicinal preparation has demonstrated its curative potential to the highest extent in case of availability of an aggravated state of numerous vital systems of the human organism.
It is a well-known fact the the destructive effect of the immune deficiency virus on the immune system starts immediately upon the moment of infection. Sensitivity to antiretroviral preparations, resistance, and capability of damaging cells are very variable among virus isolates. Numerous research workers are skeptical about the possibility of HIV elimination by using only antiretroviral preparations, probably except a short time period immediately after infection. Even if preparations result in a substantial reduction of the viral loading, the amount of lymphocyte helpers seldom comes back to the normal level. On the contrary, it usually remains over a plateau of 100 to 150/mm.sup.3. In addition, there may be grave and unforeseen consequences of the long-term usage of antiretroviral therapy, which fact requires effort aimed at development of "natural", immunity-based control of HIV infection.
Although numerous studies are carried out/aimed at support and restoration of immune competence with the use of immune modulators, cytokins, growth factors, and specific immune intervention (passive immunization and vaccination), no licensed preparations are available that could restore and support immune system functions.
The progress of AIDS involves two sides:
infection with HIV, its reproduction inside organism, and attack on patient's immune system, and PA1 increase in the activity of immune response of patient's organism, aimed at neutralizing the HIV attack, exhaustion of the functional activity of the immune system, and subsequent decrease in the production of important components of the immune system as a result of exhaustion of the leukocytic branch of hemopoiesis. PA1 suppression of HIV reproduction with the use of antiretroviral preparations, and PA1 strengthening (in HIV-infected and carriers) and restoration (in AIDS patients) of the functional activity of immune system, and compensation of insufficient own immune response. PA1 the material that is available for transplantation in a specific situation is often non optimal for a certain patient, which fact results in selection of a recipient for the material rather than selection of a medicine for the particular patient as it is in most cases; PA1 the use of a transplantation material ex tempore does not ensure sufficient infection safety of the treatment process; PA1 transplantation requires selection of material in compliance with histocompatibility systems H suppression of an immune conflict between recipient and transplant; PA1 the total amount of cells frequently comprises the only feature of a material to be transplanted; PA1 in many cases, cell transplantation material is wasted, and cell suspensions are prepared from organs of several embryos PA1 increase in the concentration of nondifferentiated lymphoid elements, including progenitor cells, PA1 guaranteed preservation of their viability and capability of producing cell posterity in the patient's organism, PA1 application of the preparation without any risk of infecting a patient with infectious diseases, PA1 absence of need to select patients on the basis of major systems of histocompatibility genes, PA1 absence of need to carry out conditioning prior to treatment, PA1 absence of need to use immune depressants, PA1 absence of "host-transplant" immune conflict and rejection reaction, PA1 application immediately upon clinical prescription, PA1 repeated and multiple treatment sessions with the use of the inventive method of treatment and inventive medicinal preparations, PA1 application, in the on-going treatment course, of the same fetal material that has been already used for this patient, PA1 economical use of fetal material in fractional doses during patient's life as the clinical need arises, PA1 unlimited storage term, PA1 transportation to substantial distances. PA1 stage 1: from room temperature down to -4.degree. C., at a rate of 1.degree. C./min; PA1 stage 2: from -4 down to -10.degree. C., at a rate of 0.1.degree. C./min; PA1 stage 3: from -10.degree. C. down to -190.degree. C., at a rate of 7.degree. C./min. PA1 total amount (contents) of nucleated cells in 1 ml (cell analyzer or visually, under microscope, in a counting chamber); PA1 Colony-forming units of the granulocyte/macrophage (CFU GM) 1 ml by methods of CFU cloning in methyl cellulose; PA1 Colony-forming units of the granulocyte, erythrocyte, monocyte/macrophage, megakaryocyte (CFU GEMM)" in 1 ml; PA1 Progenitor cells (CD34). CD34 in 1 ml (by indirect immunofluorescent test with the panel of monoclonal antibodies). PA1 a) the contents of nucleated cells is 5 to 200'10.sup.6 /ml; PA1 b) the contents of colony-forming units of granulocyte/macrophage is 20 to 200'10.sup.3 /ml; PA1 c) the contents of colony-forming units of granulocyte, erythrocyte, monocyte/macrophage, megakaryocyte is 0.5 to 50'10.sup.3 / ml; and PA1 d) the contents of progenitor cells (CD34) is 1 to 20'10.sup.6 /ml. PA1 Medicinal preparation can be administered intravenously, in drops, in the composition of 100-150 ml of isotonic solution of sodium chloride, at a rate of 20 to 40 drops a minute.
Therefore, treatment of the disease is directed at both sides of the process:
At present, much success has been achieved in the suppression of HIV replication. As you know, however, this does not result in the recovery of patients suffering from AIDS and HIV-infection.
Restoration of the functional activity of immune system constitutes an independent task in the course of treatment of patients suffering from AIDS and HIV-infection.
We suppose that the most success will be attained through combination of using antiviral therapeutic means aimed at suppression of virus replication, and the inventive medicinal preparation directed at restoration of the functional activity of the patient's immune system.
In recent years, a striking progress has taken place in the studies and application of human fetal cadaver tissues. A completely new area of therapy is being developed, i.e. cell therapy that permits to fill an insufficient functional activity of damaged and sore tissues through the application of medicinal preparations based on fetal medicinal preparation and prepared from fetal tissues.
By using medicinal preparation prepared from fetal embryo tissues, it is possible to attain such conditions where embryo cells administered to the organism of a patient may survive, locate their respective target organs and produce descendants, thereby filling deficient functional units of organs and tissues.
Embryo cells are often capable of migrating, establishing intercellular links, proliferating, differentiating, and responding to effects. They can produce considerable amounts of biologically active substances, e.g. hemopoictic growth factors, interleukins, nerve growth factor, allogenic and neurotrophic factors etc.
Embryo cells cause a weaker immune response than mature cells due to the late expression of the main antigens in the process of their maturation. Embryo cell suspensions contain much lower amounts of highly active cells such as leukocytes, endothelium of vessels, dendritic cells etc. During the early fetal period transplants do not have mature lymphocytes, and are tolerant to recipient's tissues.
Embryo cell suspensions possess higher resistance than mature cells, they are capable of surviving under lower levels of oxygen contents. Since they do not have either long processes or strong intercellular adhesion, such cells are less susceptible to traumatic damage during suspension preparation. Finally, they pass the conditions of programmed cryofreezing and unfreezing in an easier and simpler way, while maintaining all of their amazing properties and being less damaged than mature cells.
At present, applied are suspensions prepared from fetal brain, bone marrow, liver, spleen, thymus, pancreas, and culocutaneous graft.
The most successful initial attempts to apply cell suspensions involved human fetal liver and spleen.
Embryo cell suspension prepared from liver and not subjected to selection and cell grading comprises a complicated concentrated multifunctional suspension consisting of a liquid, biologically active substances and cells that are changing in a very dynamic way, depending on embryo's age.
In 1973, suspension of native cells of a fetal liver of a 7-week gestation was prepared for the first time; administration of this suspension resulted in the recovery of hemopoiesis in a patient suffering from aplastic anemia (Kelemen E. Second J. Gematol., 1973, v. 10, No.4, pp.305-308 [6]).
In recent years, by varying methods of preparing of transplantat and procedures of their application, research workers managed to achieve positive results of treating primary and secondary myelodepressive states. These preparations have been described in particular in [8, 9].
An excellent paper presented by Baechelta R. et al in I. Clin. Invest., 1993, v.91, March, pp.1067-1078 [1], shows end results of treating patients suffering from grave combined immune deficiency; here, not only immunity indices have been recovered in patients, but also availability of split chimerism and emergence of tolerance to both host and donor antigens have been demonstrated.
Another area of clinical application of fetal liver cell suspension administration comprised treatment of immunity disorders and inborn errors of metabolism; here, the most considerable experience has been accumulated by Touraine J. [11-13]. J. L. Touraine is using Fetal liver transplantation. The material was taken directly from one or several fetus corps. The only characteristic of administered material comprised the amount of administered cells. J. L. Touraine did not use any special characteristics of the administered material other than identification, in some cases, of histocompatibility antigens.
It is also clear from the paper by T. Izzi and contributors from the J. Lucarelli clinic [9], that preparation of the Fetal Liver transplant took place several hours before transplantation. Transplantats characterize of high amount of cells (10**9).
T. Izzi et al. always select material and recipient in compliance with histocompatibility antigens, which imposes a considerable restriction on their procedure.
Referring to publications by the authors working in the area of fetal liver transplantation, particularly J. Lucarelli, Touraine J. L., and others, it should be noted that they achieved much success in the treatment of patients by working within the constraints intrinsic in transplantation methods, namely:
The main object of the invention.
The object of the invention comprised development of a medicinal preparation (agent) that would combine substantial advantages of transplantation methods, and particularly would ensure transplantation of living cells having a capability to survive, reproduce and specialize inside patient's body, and advantages provided by application of medicinal preparations, and in particular would ensure convenient, safe and repeated application without patient's conditioning, and selection of a transplant for a patient (patient for transplant) in compliance with histocompatibility genes.
Full description of the acting source of the inventive medicinal preparation, taking into account its polypotent properties, polymorphous organization, features of evolution inside the organism in the course of specific differentiation, reproduction and life cycle and with account of disease specific nature and stage, is considered to be quite impossible at the modern level of knowledge, not to speak about qualitative assessment.
The inventive medicinal preparation should possess the following major advantages over the transplantation technology utilizing living cells:
A fuller understanding of the nature of the invention will be had from the following detailed description of embodiments thereof, taken in conjunction with specific Examples.